Glomerular dysfunction is a common basis for the development of chronic kidney disease (CKD), a condition with significant comorbidities and mortalities. Recent studies highlighted the role of podocyte actin cytoskeleton and the slit diaphragm in the maintenance of the glomerular filtration barrier (GFB), and the development of albuminuria (AU) and focal segmental glomerulosclerosis (FSGS). The key role of cytosolic calcium ([Ca2+]i) signaling in podocyte function and the above pathologies is established, however our mechanistic understanding of podocyte [Ca2+]i dynamics is limited to a few players (angiotensin II, TRPC5/6). Most P2 purinoceptors including P2Y2 that bind extracellular ATP signal via [Ca2+]i and have been implicated in a variety of (patho)physiological functions in many organs including paracrine cell-to-cell crosstalk, tissue injury and sclerosis. While the important function of the P2Y2 receptor in renal tubular transport and blood flow autoregulation is well established, its role in podocytes/glomerulus is largely unknown. Here we propose to study podocyte [Ca2+]i dynamics in vivo in the intact kidney in health and disease and to characterize the role and therapeutic relevance of a novel [Ca2+]i signaling mechanism in podocytes that is mediated by P2Y2 purinergic receptors. We hypothesize that P2Y2-mediated elevations in [Ca2+]i is a robust and key novel mechanism in primary podocyte injury, and also that its cell-to-cell propagation between podocytes results in amplified focal segmental dysfunction of the GFB and the development of AU and FSGS. The overall theme, to study the role of P2Y2 in podocyte/GFB injury and the effect of P2Y2 blockade on AU and FSGS, will be addressed by applying a novel imaging approach that employs intravital multiphoton microscopy (MPM) combined with new mouse models of fluorescent podocyte labeling and tagging. These studies may potentially change the current view of the pathogenesis of glomerular disorders and may also provide a clinically and immediately available, novel therapeutic approach for glomerular kidney diseases. The specific aims are to: (1) Characterize the role of P2Y2 signaling in podocyte [Ca2+]i dynamics and GFB/glomerular function in response to podocyte injury/disease. (2) Establish the amplifier function of P2Y2 signaling in the propagation of podocyte injury. (3) Test whether P2Y2 blockade can blunt/reduce the development of podocyte injury, AU and FSGS.